PKG is a serine/threonine specific protein kinase that is activated upon the intracellular generation of 3′,5′ cyclic guanosine monophosphate (cGMP) by two main types of guanylyl cyclases (GC): soluble and membrane associated Garbers, D. L., Cell 71, 1-4 (1992)). Soluble GC acts downstream of NO, while the membrane associated GC is activated through the extracellular binding of natriuretic peptides (NP).
Tyrosine nitration is a selective process as not all tyrosine residues in a protein undergo nitration under patho-physiological conditions (Ischiropoulos, H., Biochemical and Biophysical Research Communications 305, 776-783 (2003)). PKG-1α has 21 tyrosine residues in its monomeric structure, of which 9 tyrosines are located in the regulatory domain and 12 are part of the catalytic domain. In vitro and in vivo studies have demonstrated that the nitration of cGMP-dependent protein kinase G-1α (PKG-1α) is an important posttranslational event responsible for the impaired PKG activity in the lungs of acute and chronic pulmonary hypertensive lambs (Aggarwal, S., et al., Journal of Cellular Physiology (2011)), mice with hypoxia-induced pulmonary hypertension (Negash, et al., American Journal of Physiology 293, L1012-1020 (2007)), and humans with idiopathic pulmonary arterial hypertension (Zhao, et al., The Journal of Clinical Investigation 119, 2009-2018 (2009)), however, the nature and location of this nitration along the PKG-1α protein is not known.
Accordingly, there remains a need for the identification and characterization of the tyrosine residues of PKG-1α that leads to pathological and disease states, and compositions and methods for detection thereof in biological samples.
Therefore, it is an object of the invention to provide the one or more tyrosine residues of PKG-1α that can be nitrated under physiological conditions.
It is another object of the invention to provide compositions and methods for the detection of these nitrated residues in a biological sample.
It is a further object of the invention to provide methods of use thereof for assisting in the diagnosis of diseases and disorders associated with nitration of PKG-1α.